Purification and Properties of an Extracellular Endo-1, 4-β-Xylanase from Penicillium citrinum and Characterization of the Encoding Gene(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)

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An extracellular endo-1, 4-β-xylanase was purified from the culture filtrate of a filamentous fungus Penicillium citrinum FERM P-15944 grown on birch-wood xylan. The purified enzyme showed a single band on SDS-PAGE with an apparent M_r of 20, 000 and had an isoelectric point below 3.5. Xylanase activity was optimal at pH 5.0 and 55℃. The genomic DNA and cDNAs encoding this protein were cloned and sequenced. Southern blot analysis indicated that the xylanase gene (xynA) was present as a single copy in the genome. An open reading frame of 657 bp was interrupted by two introns of 65 and 55 bp, and encoded a presumed prepropeptide of 27 amino acids and a mature protein of 190 amino acids. Three distinct transcription start points were observed at positions -20 (A), -31 (A), and -36 (A) from the start codon. The 5´-noncoding region had a putative TATA box at nt -66 (TATAAA). The xynA cDNA was functionally expressed under the control of the alcohol oxidase I gene promoter in the methylotrophic yeast Pichia pastoris. A neighbor-joining tree showed that the P. citrinum enzyme is closely related to several other fungal xylanases belonging to the glycoside hydrolase family 11 : Trichoderma reesei XYN2, Aspergillus niger xynNB, Penicillium funiculosum xynC, Penicillium sp. strain 40 xynA, Chaetomium gracile cgxB, and Aspergillus nidulans xlnA and xlnB.
社団法人日本生物工学会の論文
2005-12-25